Hepatitis delta virus (HDV) is a viroid-like agent causing severe forms of hepatitis in infected people. It is a naturally occurring satellite of human hepatitis B virus (HBV) and has been shown to be a human pathogen. Rizzetto, M. et al., Proc. Nat'l Acad. Sci. USA 77: 6124-6128 (1980). HDV infection may occur as a coinfection with HBV, and as a superinfection of an HBV carrier. Each type of infection may cause acute or fulminant hepatitis. Superinfection usually causes chronic hepatitis and eventual death due to cirrhosis and liver failure. Approximately 10,000 new HDV infections occur annually in the United States leading to 1,000 hospitalizations and 50 deaths due to fulminant hepatitis. Approximately 70,000 persons are chronic HDV carriers, and an estimated 750-1,000 die annually of chronic liver disease.
Hepatitis delta virions are 36-nm particles with an envelope composed of hepatitis B surface antigen. The internal part of these particles consists of a HDV-specific protein, hepatitis delta antigen ("HDAg"), and a small genomic RNA. Bonino, F., et al., Infect. Immunol. 43:1000-1005 (1986). HDAg found in the sera and liver of infected humans or experimentally infected animals has been shown to be composed of two protein species of between 24 and 30 kDa or a major 26 kDa protein and multiple minor proteins. Bonino, F., et al., J. Virol. 58: 945-50 (1986) and Puig, J., et al., in: Hepadna Viruses (Robinson, W. et al. eds.) Alan R. Liss, N.Y. pp. 579-590 (1987). All of these proteins appear to be related in sequence and are encoded by the large open reading frame (ORF5) of the HDV antigenomic RNA. Makino, S. et al., Nature 329:343-346 (1987) and Wang, E.-S., et al., Nature, 323:508-513 (1986).
European patent application no. 251 575 by Chiron Corporation entitled "Hepatitis Delta Diagnostics and Vaccines, Their Preparation and Use" (published Jan. 7, 1988), discloses the entire genome of the hepatitis delta virus and several open reading frames in both the genomic and complementary strands. One open reading frame, ORF5, encodes two viral polypeptides, p24 and p27. The ORF5 products are considered to be useful in HDV diagnosis and vaccines.
International Patent Application No. PCT/US90/06077by Lemon, entitled "Immunological Domains Of Hepatitis Delta Virus Antigen" (published May 16, 1991 as World Patent No. 91/06562), discloses the synthesis of 209 overlapping hexapeptides spanning the 214 amino acid residues of the protein HDAg (ORF2 expression product) in order to map its antigenic domains. Domains recognized by antibodies present in serum from human chronic carriers of HDV included residues 2-7, 63-74, 86-91, 94-100, 159-172, 174-195 and 107-207. Also, residues 2-17, 156-184 and 197-211 were synthesized in bulk and were found to possess significant antigenic activity by microtiter ELISA. It was concluded that the entire 214 amino acids of HDAg are expressed during infection in vivo and that the aforementioned peptides would be useful as diagnostic agents and as vaccines.
Khudyakov, et al., "Amino Acid Sequence Similarity Between The Terminal Protein Of Hepatitis B Virus And Predicted Hepatitis Delta Virus Gene Product", FEBS 262(2): 345-348 (March 1990), identifies HDAg' as a new HDV protein. They also propose that HDAg' is encoded by HDAg mRNA, is synthesized by a translation slippage mechanism and may be involved with interfering in Hepatitis B virus replication. FIG. 2, on page 346, presents the genetic organization of the region 631-730 of HDV antigenomic RNA strand. It also shows the sequences for the translation products of the HDAg' open reading frame and indicates probable slippage sites in translation.
Thus far, hepatitis delta virus infection has been diagnosed by the detection of HDAg, or by the detection of antibody to the antigen (anti-HD). But the pathological significance and diagnostic value of HDAg' as a marker has not been determined and such an assessment is needed. The use of HDAg' polypeptides in vaccines against HDV infection has also previously not been investigated.